High‐resolution proteomic and lipidomic analysis of exosomes and microvesicles from different cell sources

Abstract

Extracellular vesicles (EVs), including exosomes and microvesicles (MVs), are explored for use in diagnostics, therapeutics and drug delivery. However, little is known about the relationship of protein and lipid composition of EVs and their source cells. Here, we report high-resolution lipidomic and proteomic analyses of exosomes and MVs derived by differential ultracentrifugation from 3 different cell types: U87 glioblastoma cells, Huh7 hepatocellular carcinoma cells and human bone marrow-derived mesenchymal stem cells (MSCs). We identified 3,532 proteins and 1,961 lipid species in the screen. Exosomes differed from MVs in several different areas: (a) The protein patterns of exosomes were more likely different from their cells of origin than were the protein patterns of MVs; (b) The proteomes of U87 and Huh7 exosomes were similar to each other but different from the proteomes of MSC exosomes, whereas the lipidomes of Huh7 and MSC exosomes were similar to each other but different from the lipidomes of U87 exosomes; (c) exosomes exhibited proteins of extracellular matrix, heparin-binding, receptors, immune response and cell adhesion functions, whereas MVs were enriched in endoplasmic reticulum, proteasome and mitochondrial proteins. Exosomes and MVs also differed in their types of lipid contents. Enrichment in glycolipids and free fatty acids characterized exosomes, whereas enrichment in ceramides and sphingomyelins characterized MVs. Furthermore, Huh7 and MSC exosomes were specifically enriched in cardiolipins; U87 exosomes were enriched in sphingomyelins. This study comprehensively analyses the protein and lipid composition of exosomes, MVs and source cells in 3 different cell types.

Matrix vesicles from chondrocytes and osteoblasts: Their biogenesis, properties, functions and biomimetic models

“…The majority of the purification methods do not yield a single type of EVs [67]. This is also true for the purification of MVs.…”

The biology of extracellular microvesicles

“…MVs display unique lipid characteristics, such as the externalization of the phospholipid phosphatidylserine (PS), which has been demonstrated to promote uptake by recipient cells . The phospholipid lysophosphatidylcholines, the sphingolipid sphingomyelins, and acylcarnitines, the fatty acyl esters of L‐carnitine, have also been found to be enriched in MVs . The prominent plasma membrane lipid cholesterol is thought to play an important role in MV formation, as its depletion reduces MV formation .…”

“…The cargoes contained within EVs reflect both the intracellular origin of the cargo as well as the cell type from which the vesicle was derived. We refer readers to the many profiling studies which have been published on this subject for further information . Prototypical exosome markers include tetraspannins such as CD9, CD63 and CD81, and ESCRT proteins Alix and TSG101 .…”

Thematic Review Series: Exosomes and Microvesicles: Lipids as Key Components of their Biogenesis and Functions Extracellular vesicles and their content in bioactive lipid mediators: more than a sack of microRNA

“…For instance, transmembrane proteins such as tetraspanins (CD9, CD63, CD81), and tumor susceptibility gene 101 (TGS101) and Alix, accessory molecules from the endosomal sorting complex, are mostly associated with exosomes (10). Conversely, proteins of organelle origin such as those from the endoplasmic reticulum, Golgi, mitochondria, or nucleus, are preferentially found in microvesicles and are rarely found in exosomes (10,11).…”

“…Extracellular vesicles imperatively comprise a lipid moiety, and their cholesterol, sphingomyelin, phosphatidylserine, and glycosphingolipid content is richer than their cellular sources (11)(12)(13)(14)(15)(16). The studies that originally reported their presence in blood determined that the EV membrane could support the coagulation cascade (2, 17).…”